KR102315879B1 - Novel Rhizopus oligosporus strain and antibacterial and antifungal composition comprising the same - Google Patents

Abstract

The present application relates to: a Rhizopus oligosporus CJCC02-20 strain deposited with an accession number KCCM12893P; an antibacterial composition comprising, as an active ingredient, any one or more selected from the group consisting of the strain, the spore of the strain, the culture of the strain, a concentrate thereof, and a dried product thereof; a food composition comprising the antibacterial composition; a quasi-drug composition; a health functional food composition; or a pharmaceutical composition for prevention or treatment of microbial infectious diseases caused by fungi, bacteria, or a combination thereof, by comprising the antimicrobial composition. ​Since the antibacterial composition comprising the Rhizopus oligosporus CJCC02-20 strain according to the present application has excellent antibacterial activities against fungi and bacteria, the composition can be useful as food, medicine, quasi-drug, health functional food, etc.

Description

Novel Rhizopus oligosporus strain and antibacterial composition comprising the same

The present application is a Rhizopus oligosporus CJCC02-20 (Rhizopus oligosporus CJCC02-20) strain deposited with accession number KCCM12893P; An antibacterial composition comprising, as an active ingredient, any one or more selected from the group consisting of the strain, the spore of the strain, the culture of the strain, its concentrate and its dried product; a food composition comprising the antimicrobial composition; feed composition; quasi-drug composition; health functional food composition; Or it relates to a pharmaceutical composition for the prevention or treatment of microbial infections caused by fungi, bacteria, or a combination thereof, including the antibacterial composition.

Fermented food manufactured in a traditional way is always vulnerable to fungi and food poisoning bacteria during the storage, manufacturing and distribution of raw materials. Fungi and food poisoning-causing bacteria are known to survive heat treatment and cause spoilage and spoilage of food or food poisoning.

In order to prevent spoilage and proliferation of food poisoning bacteria, and to maintain the quality of the food for a long time, a lot of research on antibacterial substances has been conducted. Among antibacterial substances, the need for research on natural preservatives with excellent safety and economical efficiency is increasing.

As a representative natural preservative, bacteriocin refers to a natural antibacterial protein or protein-based material produced by microorganisms. Bacteosin is a protein, so it can be decomposed in the human body, and thus its safety has been proven. In contrast to the secondary metabolites of existing antibiotics that cause side effects in the human body, the need for research on bacteriocins is increasing. With respect to the bacteriocin, studies on lactic acid bacteria-derived bacteriocins have been predominant until recently. Lactobacillus-derived bacteriocin has the advantage of being proven safe with a GRAS grade, but it is not widely used because the antibacterial activity range is specific to Gram-positive bacteria.

With respect to the antimicrobial properties of Rhizopus oligosporus , antibiotic peptide (ABP) and Linoleic acid, which are antibacterial substances produced from the culture medium of Rhizopus oligosporus , are known to control the gram-positive genus Straphyloccoccus and Bascillus (Kusumah et al. , 2020; Nowak and Steinkraus, 1988; Roubos-van den Hil et al., 2010; Wang et al., 1969). However, only antibacterial activity against specific microorganisms that are gram-positive has been reported, and there is no report on the effect on various strains causing decomposition or food poisoning. Furthermore, since Rhizopus oligosporus is a useful bacterium in fermented food, when the bacteriocin is applied to fermented food, it is difficult to see that it exhibits specific activity in rot-inducing fungi and food poisoning bacteria. have.

Accordingly, the present inventors made intensive research efforts to develop a method for inhibiting various fungi and bacteria while harmless to the human body. As a result, Rhizopus oligosporus CJCC02-20 newly isolated from tempe has antibacterial activity against fungi as well as Rather, the present application was completed by confirming that there is an antibacterial activity that specifically inhibits only major harmful microorganisms in food such as food poisoning bacteria.

The present application is a Rhizopus oligosporus CJCC02-20 (Rhizopus oligosporus CJCC02-20) strain deposited with accession number KCCM12893P; An antibacterial composition comprising, as an active ingredient, any one or more selected from the group consisting of the strain, the spore of the strain, the culture of the strain, its concentrate and its dried product; a food composition comprising the antimicrobial composition; feed composition; quasi-drug composition; It relates to a pharmaceutical composition for preventing or treating microbial infections caused by fungi, bacteria, or a combination thereof, including a health functional food composition or the antibacterial composition.

One object of the present application is to provide a Rhizopus oligosporus CJCC02-20 (Rhizopus oligosporus CJCC02-20) strain deposited with accession number KCCM12893P.

Another object of the present application is to include, as an active ingredient, any one or more selected from the group consisting of the Rhizopus oligosporus CJCC02-20 strain, the spore of the strain, the culture of the strain, its concentrate, and its dried product, To provide an antibacterial composition.

Another object of the present application is to provide a food composition comprising the antibacterial composition.

Another object of the present application is to provide a feed composition comprising the antibacterial composition.

Another object of the present application is to provide a quasi-drug composition comprising the antibacterial composition.

Another object of the present application is to provide a health functional food composition comprising the antibacterial composition.

Another object of the present application is to provide a pharmaceutical composition for preventing or treating microbial infections caused by fungi, bacteria, or a combination thereof, including the antibacterial composition.

In order to achieve the above object, one aspect of the present application provides a Rhizopus oligosporus CJCC02-20 strain deposited with accession number KCCM12893P.

The present inventors have identified a novel strain belonging to Rhizopus oligosporus having excellent antibacterial activity against fungi and food poisoning-causing bacteria that cause deterioration and contamination of fermented food as follows.

As a result of isolating various types of Rhizopus oligosporus from conventional tempeh and testing for fungi, yeast, etc. and food poisoning bacteria, a strain with excellent antibacterial activity was selected, and gene sequencing to identify the selected strain. Analysis was performed. The 18S rRNA base sequence of the selected strain is described in SEQ ID NO: 1, the antibiotic peptide (ABP) gene sequence is described in SEQ ID NO: 2, and the Chitinase (Chi2) gene sequence is described in SEQ ID NO: 3. Based on the above genetic analysis, the strain was identified as a Rhizopus oligosporus strain, and was named Rhizopus oligosporus CJCC02-20, and was sent to the Korea Microorganism Conservation Center (KCCM), an international depository under the Budapest Treaty, on December 09, 2020. It was deposited on the date and was given an accession number KCCM12893P.

According to one embodiment of the present application, the newly isolated Rhizopus oligosporus CJCC02-20 has excellent antibacterial activity against fungi and food poisoning-inducing bacteria, and exhibits antibacterial activity that can act simultaneously on various fungi and food poisoning-causing bacteria. (Example 3).

The fungi and bacteria may be of the genus Aspergillus , the genus Bacillus , and the genus Escherichia , but is not limited thereto.

As an embodiment of the present application, the Rhizopus oligosporus CJCC02-20 is a fungus that has antibacterial activity against Aspergillus oryzae , Aspergillus niger , or a combination thereof. However, it is not limited to the type of the fungus.

As an embodiment of the present application, the Rhizopus oligosporus CJCC02-20 may have antibacterial activity against Bacillus cereus , Escherichia coli , or a combination thereof, but depending on the type of bacteria It is not limited.

As an embodiment of the present application, the Rhizopus oligosporus CJCC02-20 may have specific antibacterial activity against Aspergillus duck material, Aspergillus niger or E. coli, but is not limited thereto.

The Aspergillus duck material and Aspergillus niger are known as strains that cause deterioration and contamination of fermented food, and Bacillus cereus and E. coli are known as food poisoning causative bacteria, and Rhizopus oligospo having excellent antibacterial activity. Russ CJCC02-20 can be used for various purposes.

As another aspect for achieving the above object, the present application relates to the novel isolated Rhizopus oligosporus CJCC02-20 strain, the spores of the strain, the culture of the strain, its concentrate, its extract, and its dried product. Any one or more selected from the group consisting of; and a carrier.

In this case, the carrier may be a naturally or non-naturally occurring carrier.

In one embodiment of the present application, the dried product may be a freeze-dried product.

In addition, the composition may further comprise a cryoprotectant. For example, one or more cryoprotectants selected from the group consisting of glycerol, trehalose, maltodextrin, powdered skim milk and starch may be further included. The cryoprotectant of the present application may be included in 0.01 wt% to 20 wt%, 0.01 wt% to 10 wt%, based on the total weight of the composition of the present application, specifically, the glycerol of the present application is 5 wt% to 20 wt%, the present application of trehalose is 2 wt% to 10 wt%, maltodextrin of the present application is 2 wt% to 10 wt%, skim milk powder of the present application is 0.5 wt% to 2 wt%, starch of the present application is 0.1 wt% to 1 wt% % may be included in the composition of the present application.

As another aspect for achieving the above object, the present application relates to the novel isolated Rhizopus oligosporus CJCC02-20 strain, the spores of the strain, the culture of the strain, its concentrate, its extract, and its dried product It provides an antibacterial composition comprising any one or more selected from the group consisting of as an active ingredient.

At this time, the state of the strain may be a liquid state or a dry state, but is not limited thereto.

The antimicrobial composition may have antibacterial activity against fungi, bacteria, or a combination thereof, and specifically, may have specific antibacterial activity against Aspergillus duck material, Aspergillus niger or E. coli, and Bacillus cerebellum. It may have an enhanced antibacterial activity in the mouse, but is not limited thereto.

As used herein, the term “spore” refers to a propagation cell such as bacteria, and for the purpose of the present application, the spore refers to a propagation cell of Rhizopus oligosporus CJCC02-20.

In the present application, the spores of the Rhizopus oligosporus CJCC02-20 exhibit antibacterial activity against fungi and food poisoning bacteria, and thus can be used in foods, feeds, pharmaceuticals, quasi-drugs, and health functional foods having antibacterial activity.

As used herein, the term "culture" refers to a product obtained after culturing the strain, and may be a culture stock solution including cells, or may be cells obtained by removing or concentrating cultured strains and culture supernatant. The composition of the culture may additionally include not only components necessary for culturing Rhizopus oligosporus, but also components that act synergistically on the growth of Rhizopus oligosporus, and the composition according to the conventional technique in the art It can be easily selected by those who have it.

As used herein, the term "concentrate" refers to the concentration of the culture.

As used herein, the term "extract" refers to an extract from the culture or its concentrate, and is not limited thereto, as long as it is an extract capable of exhibiting a specific antibacterial effect on fungi and bacteria of the present application, but is not limited thereto. of the diluted or concentrated solution, the dried product obtained by drying the extract, or these crude products or purified products, and all fractions obtained by fractionation thereof.

As used herein, the term “dried product” means drying the culture, its concentrate, its extract, and its fraction. The drying method may include, but is not limited to, ventilation drying, natural drying, spray drying and freeze drying.

The newly isolated Rhizopus oligosporus CJCC02-20 of the present application may be cultured through a conventional culturing method of the Rhizopus oligosporus strain, but is not limited thereto. As the medium, a natural medium or a synthetic medium may be used. As a carbon source of the medium, for example, glucose, sucrose, dextrin, glycerol, starch, etc. may be used, and as a nitrogen source, peptone, meat extract, yeast extract, dried yeast, soybean, ammonium salt, nitrate and other organic or inorganic Nitrogen containing compounds may be used, but are not limited to these components. As the inorganic salt contained in the medium, magnesium, manganese, calcium, iron, phosphorus, etc. may be used, but is not limited thereto. In addition to the components of the carbon source, nitrogen source and inorganic salt, amino acids, vitamins, nucleic acids and related compounds may be added to the medium.

The antibacterial composition may include other by-products generated in the process of culturing Rhizopus oligosporus CJCC02-20 together.

As another aspect for achieving the above object, the present application provides a food composition comprising the antimicrobial composition.

The type of the food is not particularly limited, and may include all foods in a conventional sense. Non-limiting examples of foods to which the above substances can be added include soy sauce, fermented foods including salted fish, sauces, meat, sausage, bread, chocolate, candies, snacks, confectionery, pizza, ramen, other noodles, gums, ice cream Dairy products including dairy products, various soups, beverages, tea, drinks, alcoholic beverages, and vitamin complexes are included. When the composition is used as a food additive, the composition may be added as it is or used together with other foods or food ingredients, and may be appropriately used according to a conventional method.

Specifically, the food composition may be a fermented food, specifically soy sauce, salted fish, and more specifically tempe, but is not limited thereto.

In addition, the food composition may be a sauce containing tempeh, but is not limited thereto.

The food composition may include a food pharmaceutically acceptable carrier.

The type of the carrier is not particularly limited, and any carrier commonly used in the art may be used.

In addition, the food composition may include additional ingredients that are commonly used in food compositions to improve odor, taste, and vision. For example, vitamins A, C, D, E, B1, B2, B6, B12, niacin, biotin, folate, pantothenic acid, and the like may be included. In addition, minerals such as zinc (Zn), iron (Fe), calcium (Ca), chromium (Cr), magnesium (Mg), manganese (Mn), copper (Cu), chromium (Cr); and amino acids such as lysine, tryptophan, cysteine, and valine.

In addition, the food composition includes a preservative (potassium sorbate, sodium benzoate, salicylic acid, sodium dehydroacetate, etc.), a disinfectant (bleaching powder and high bleaching powder, sodium hypochlorite, etc.), an antioxidant (butylhydroxyanisole (BHA), butyl hydro Loxytoluene (BHT), etc.), coloring agents (tar pigments, etc.), coloring agents (sodium nitrite, sodium nitrite, etc.), bleach (sodium sulfite), seasonings (MSG sodium glutamate, etc.), sweeteners (dulcin, cyclamate, saccharin, etc.) , sodium, etc.), fragrances (vanillin, lactones, etc.), swelling agents (alum, D-potassium hydrogen tartrate, etc.), strengthening agents, emulsifiers, thickeners (flavors), filming agents, gum base agents, foam inhibitors, solvents, improving agents, dispersing agents, etc. of food additives. The additive may be selected according to the type of food and used in an appropriate amount.

In particular, the present inventors found that the newly isolated Rhizopus oligosporus CJCC02-20 of the present application is an enteric fermented mold, Aspergillus duck material, Aspergillus niger, a food contamination source, Bacillus cereus, a food poisoning bacterium, and enteropathogenic bacteria. It was confirmed that there is an effect of simultaneously inhibiting E. coli (Example 3, Fig. 1), and as a result of applying the antibacterial composition to tempeh, a fermented food, it was confirmed that it was possible to maintain the effect of controlling food poisoning bacteria while implementing the traditional flavor ( Example 4), it can be used directly for all foods, including fermented foods.

As another aspect for achieving the above object, the present application provides a pharmaceutical composition for preventing or treating a microbial infection, including the antibacterial composition.

As used herein, the term "prevention" may refer to any action of inhibiting or delaying the onset of a microbial infectious disease by administering the composition for preventing or treating a microbial infection disease according to the present application to an individual.

As used in the present application, the term "treatment" may refer to any act of administering the composition of the present application to a subject suspected of having a microbial infection disease to improve or benefit from symptoms of a microbial infection disease.

As used herein, the term “improvement” may refer to any action that at least reduces a parameter related to a treated condition, for example, the degree of a symptom.

As used herein, the term "individual" may mean any animal, including humans, that has or is likely to develop a microbial infection disease.

In this case, the microbial infection disease is a disease caused by infection by microorganisms, preferably a disease caused by infection by fungi or bacteria.

As an embodiment of the present application, the fungus may be Aspergillus duck material, Aspergillus niger, or a combination thereof, but is not limited thereto.

In one embodiment of the present application, the bacteria may be Bacillus cereus, E. coli, or a combination thereof, but is not limited thereto.

In addition, as an embodiment of the present application, the microbial infection disease may be any one or more selected from the group consisting of food poisoning, diarrhea, vomiting, enteritis, gastroenteritis, constipation, abdominal pain and abdominal distension, but is not limited thereto.

In one embodiment of the present application, the enhanced antibacterial activity against Bacillus cereus, a bacterium that can cause food poisoning, and specific antibacterial activity against enteropathogenic E. coli were confirmed, and the fungi Aspergillus duckjae and Aspergillus As the specific antifungal activity against Niger was confirmed, the antibacterial composition of the present application can be effectively used in a pharmaceutical composition for preventing or treating microbial infectious diseases.

The composition for preventing or treating a microbial infection according to the present application may further include a pharmaceutically acceptable carrier, and is formulated with the carrier to provide food, health functional food, pharmaceuticals, quasi-drugs, feed additives and drinking water additives, etc. can be provided as

As used herein, the term “pharmaceutically acceptable carrier” may refer to a carrier or diluent that does not inhibit the biological activity and properties of an administered compound without irritating the organism.

The type of carrier usable in the present application is not particularly limited, and any carrier commonly used in the art and pharmaceutically acceptable may be used. Non-limiting examples of the carrier include saline, sterile water, Ringer's solution, buffered saline, albumin injection solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and the like. These may be used alone or in mixture of two or more.

In addition, if necessary, other conventional additives such as antioxidants, buffers and/or bacteriostats can be added and used, and diluents, dispersants, surfactants, binders, lubricants, etc. It can be used by formulating it into a dosage form, pill, capsule, granule, or tablet.

The method of administering the composition for preventing or treating microbial infection according to the present application is not particularly limited, and may follow a method commonly used in the art. As a non-limiting example of the administration method, the composition may be administered by oral administration or parenteral administration. The composition for preventing, improving, or treating the microbial infectious disease of the present application may be prepared in various formulations according to the desired administration method.

The pharmaceutical dosage form of the antimicrobial substance of the present application may be used in the form of a pharmaceutically acceptable salt thereof, and may be used alone or in combination with other pharmaceutically active compounds as well as in an appropriate group.

As another aspect, the present application provides a method for preventing or treating a microbial infectious disease comprising administering to an individual a composition for preventing or treating the microbial infectious disease. As used herein, the term "individual" may mean any animal, including humans, that has or is likely to develop a microbial infection disease.

Specifically, the prevention or treatment method of the present application may include administering the composition in a pharmaceutically effective amount to an individual having or at risk of developing a microbial infection disease.

A suitable total daily amount of the composition containing the extract may be determined by the treating physician within the scope of correct medical judgment, and may be administered once or divided into several doses. However, for the purposes of the present application, a specific therapeutically effective amount for a particular patient depends on the type and extent of the response to be achieved, the specific composition, including whether other agents are used in some cases, the patient's age, weight, general health, It is preferable to apply differently depending on various factors including sex and diet, administration time, administration route and secretion rate of the composition, treatment period, drugs used together or concurrently with a specific composition, and similar factors well known in the pharmaceutical field.

The antimicrobial substance of the present application may be administered to mammals such as rats, mice, livestock, and humans by various routes. Any mode of administration can be envisaged, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection.

As another aspect, the present application provides a health functional food for preventing or improving microbial infectious diseases comprising the antibacterial composition.

At this time, the meaning of microbial infection disease, prevention, and improvement is as described above.

As used herein, the term "health functional food (health functional food or nutraceutical)" is the same term as food for specific health, functional food, and health food. It means a food with high effect, and the food may be prepared in various forms such as tablets, capsules, powders, granules, liquids, pills, etc. to obtain a useful effect for preventing or improving hair loss-related diseases.

The health functional food may include a food-logically acceptable carrier, which is as described above.

The health functional food of the present application can be manufactured by a method commonly used in the art, and at the time of manufacture, it can be prepared by adding raw materials and components commonly added in the art. In addition, if the formulation of the health functional food is also recognized as a health functional food, it may be manufactured without limitation. The health functional food of the present application can be manufactured in various forms, and unlike general drugs, it has the advantage that there are no side effects that may occur during long-term administration of the drug by using food as a raw material, and has excellent portability. health functional food can be consumed as a supplement to enhance the prevention or improvement effect of microbial infections.

Specifically, the food is a food prepared by adding the antimicrobial composition to food materials such as beverages, teas, spices, gum, and confectionery, or encapsulating, powdering, suspension, etc., for example, various foods, beverages, gum , tea, vitamin complex, health functional food, etc.

The food can be prepared in dosage forms such as tablets, granules, powders, capsules, liquid solutions and pills according to a known manufacturing method, and the content of the composition according to the present application can be adjusted according to the dosage form. There is no particular limitation on other ingredients other than containing the antimicrobial composition according to the present application as an active ingredient, and various conventional flavoring agents or natural carbohydrates may be included as additional ingredients.

As another aspect, the present application provides a quasi-drug composition comprising the antibacterial composition.

In the present application, the term "quasi-drug" refers to fibers, rubber products, or the like, which are used for the purpose of treating, alleviating, treating or preventing diseases of humans or animals, have weak action on the human body, or do not directly act on the human body. or non-machines and the like, as an article corresponding to one of the preparations used for sterilization, insecticide and similar purposes for the prevention of infection type, for the purpose of diagnosing, treating, alleviating, treating or preventing diseases of humans or animals It can refer to items other than instruments, machines, or devices, and items used for the purpose of pharmacologically affecting the structure and function of humans or animals, excluding those that are not instruments, machines, or devices. In addition, the quasi-drugs may include external preparations for skin and personal care products. Preferably, it may be a disinfectant cleaner, shower foam, gargrin, wet tissue, detergent soap, hand wash, or ointment, but is not limited thereto.

The quasi-drug composition may include a pharmaceutically acceptable carrier, as described above.

The present inventors have confirmed that the newly isolated Rhizopus oligosporus CJCC02-20 has an antibacterial effect on the fungi and bacteria, and the antibacterial composition using the novel strain can be effectively used for quasi-drugs.

As another aspect for achieving the above object, the present application provides a feed composition comprising the antibacterial composition.

As used herein, the term “feed” may refer to any natural or artificial diet, one meal, etc., or a component of the one meal meal, for or suitable for the animal to eat, ingest, and digest.

The type of feed is not particularly limited, and feed commonly used in the art may be used. The composition for feed may include a feed additive. The feed additive of the present application corresponds to an auxiliary feed under the Feed Management Act, and may include probiotics. Non-limiting examples of the feed include plant feeds such as grains, root fruits, food processing by-products, algae, fibers, pharmaceutical by-products, oils and fats, starches, gourds or grain by-products; and animal feeds such as proteins, inorganic materials, oils and fats, minerals, oils and fats, single cell proteins, zooplankton or food. These may be used alone or in mixture of two or more.

In this case, the feed composition including the antimicrobial composition of the present application may further include an excipient, a diluent, and an additive.

It was confirmed that the newly isolated Rhizopus oligosporus CJCC02-20 of the present application has the effect of simultaneously inhibiting the fungi Aspergillus duckjae, Aspergillus niger and the food poisoning bacteria Bacillus cereus and Escherichia coli. , The present application provides a feed composition having antibacterial and antifungal activity, thereby preventing microbial infection diseases including food poisoning of livestock.

As another aspect for achieving the above object, the present application provides a method for producing an antibacterial composition, comprising the step of culturing the newly isolated Rhizopus oligosporus CJCC02-20 in a medium.

In the present application, the term "cultivation" means growing the Rhizopus oligosporus CJCC02-20 of the present application under moderately controlled environmental conditions. The culturing process of the present application may be made according to a suitable medium and culture conditions known in the art. Such a culture process can be easily adjusted and used by those skilled in the art according to the selected strain. Specifically, the culture may be a batch, continuous and/or fed-batch, but is not limited thereto.

As used herein, the term "medium" refers to a material in which nutrients required for culturing Rhizopus oligosporus CJCC02-20 of the present application are mixed as a main component, and nutrients including water essential for survival and development and growth factors. Specifically, the medium and other culture conditions used for culturing the Rhizopus oligosporus CJCC02-20 of the present application may be used without any particular limitation as long as the medium used for culturing conventional microorganisms, but the Rhizopus oligo of the present application Sporus CJCC02-20 can be cultured under aerobic conditions while controlling temperature, pH, etc. in a normal medium containing an appropriate carbon source, nitrogen source, phosphorus, inorganic compound, amino acid and/or vitamin.

As the carbon source in the present application, carbohydrates such as glucose, saccharose, lactose, fructose, sucrose, maltose; sugar alcohols such as mannitol and sorbitol; organic acids such as pyruvic acid, lactic acid, citric acid and the like; Amino acids such as glutamic acid, methionine, lysine, and the like may be included. In addition, natural organic nutrient sources such as starch hydrolyzate, molasses, blackstrap molasses, rice winter, cassava, sugar cane offal and corn steep liquor can be used, specifically glucose and sterilized pre-treated molasses (i.e., converted to reducing sugar). molasses) may be used, and other appropriate amounts of carbon sources may be variously used without limitation. These carbon sources may be used alone or in combination of two or more, but is not limited thereto.

Examples of the nitrogen source include inorganic nitrogen sources such as ammonia, ammonium sulfate, ammonium chloride, ammonium acetate, ammonium phosphate, anmonium carbonate, and ammonium nitrate; Amino acids such as glutamic acid, methionine, glutamine, and organic nitrogen sources such as peptone, NZ-amine, meat extract, yeast extract, malt extract, corn steep liquor, casein hydrolyzate, fish or degradation products thereof, defatted soybean cake or degradation products thereof, etc. can be used These nitrogen sources may be used alone or in combination of two or more, but is not limited thereto.

The phosphorus may include potassium first potassium phosphate, second potassium phosphate, or a sodium-containing salt corresponding thereto. As the inorganic compound, sodium chloride, calcium chloride, iron chloride, magnesium sulfate, iron sulfate, manganese sulfate, calcium carbonate, etc. may be used, and in addition, amino acids, vitamins and/or appropriate precursors may be included. These components or precursors may be added to the medium either batchwise or continuously. However, the present invention is not limited thereto.

In addition, during the culture of Rhizopus oligosporus CJCC02-20 of the present application, compounds such as ammonium hydroxide, potassium hydroxide, ammonia, phosphoric acid, sulfuric acid, etc. may be added to the medium in an appropriate manner to adjust the pH of the medium. In addition, during culturing, an antifoaming agent such as fatty acid polyglycol ester may be used to suppress bubble formation. In addition, in order to maintain the aerobic state of the medium, oxygen or oxygen-containing gas may be injected into the medium, or nitrogen, hydrogen or carbon dioxide gas may be injected without injection of gas or without injection of gas to maintain anaerobic and microaerobic conditions, which are limited thereto. it is not

In the culture of the present application, the culture temperature may be maintained at 20 to 45 °C, specifically, 25 to 40 °C, and may be cultured for about 10 to 160 hours, but is not limited thereto.

The method for preparing the antimicrobial composition of the present application includes the steps of preparing the Rhizopus oligosporus CJCC02-20 strain of the present application, preparing a medium for culturing the strain, or a combination thereof (regardless of the order, in any order), for example, before the culturing step, may further include.

The method for preparing the antibacterial composition of the present application may further include recovering the antimicrobial material from the culture medium (cultured medium) or the Rhizopus oligosporus CJCC02-20 strain of the present application. The recovering step may be further included after the culturing step.

At this time, the antibacterial material is included without limitation as long as it is a material having antibacterial activity produced by the Rhizopus oligosporus CJCC02-20 strain, and in one embodiment, it may be crude extracts or a compound, but is not limited thereto.

The recovery may be to collect the desired antibacterial material using a suitable method known in the art according to the culture method of the microorganism of the present application, for example, a batch, continuous or fed-batch culture method. For example, centrifugation, filtration, treatment with a crystallized protein precipitating agent (salting out method), extraction, ultrasonic disruption, ultrafiltration, dialysis, molecular sieve chromatography (gel filtration), adsorption chromatography, ion exchange chromatography, affinity Various chromatography such as island chromatography, HPLC, or a combination thereof may be used, and a desired antimicrobial substance may be recovered from the medium or microorganism using a suitable method known in the art.

In addition, the method for preparing the antimicrobial composition of the present application may additionally include a purification step. The purification may be performed using a suitable method known in the art. In one example, when the method for preparing the antimicrobial composition of the present application includes both the recovery step and the purification step, the recovery step and the purification step are performed concurrently (or continuously) regardless of the order, or simultaneously or in one step It may be integrated and performed, but is not limited thereto.

As another aspect for achieving the above object, the present application provides a food composition comprising the novel isolated Rhizopus oligosporus CJCC02-20 strain or a fermented product using the strain.

The Rhizopus oligosporus CJCC02-20 and food are as described above.

As used herein, the term "fermented product" refers to a medium obtained by inoculating and culturing the Rhizopus oligosporus CJCC02-20 strain in a medium containing the main raw material of food or a culture containing a microorganism cultured with the medium. it means.

In addition, the fermented product may be a culture obtained by inoculating and culturing the Rhizopus oligosporus CJCC02-20 strain in the food raw material. The food raw material may be grains, but is not limited thereto.

In one embodiment of the present application, the grains may be rice, barley, wheat, corn, and the like, and specifically, may be wheat, wheat bran, or rice, but is not limited thereto.

The fermented product may be comprehensively interpreted as including all of the filtrate, dilution, and concentrate thereof, purified product, dried product, pulverized product, and the like of the medium or culture, but is not limited thereto.

As another aspect for achieving the above object, the present application provides a method for producing a food comprising the step of inoculating the newly isolated strain Rhizopus oligosporus CJCC02-20.

The Rhizopus oligosporus CJCC02-20 and food are as described above.

In one embodiment of the present application, the food manufacturing method may be a manufacturing method of a fermented food comprising the step of inoculating and culturing grains with the Rhizopus oligosporus CJCC02-20. The fermented food obtained by inoculating and culturing the strain may be specifically soy sauce or salted fish, and more specifically, may be tempeh or a sauce containing tempeh, but is not limited thereto.

As another aspect for achieving the above object, the present application provides a feed composition comprising the novel isolated strain Rhizopus oligosporus CJCC02-20.

The Rhizopus oligosporus CJCC02-20 and feed are the same as described above.

Since the antibacterial composition comprising the Rhizopus oligosporus CJCC02-20 strain according to the present application has antibacterial activity against fungi and bacteria, it can be usefully used as food, medicine, quasi-drug, health functional food, etc.

1 is a result of measuring the antibacterial activity against Aspergillus duckjae, Aspergillus niger and E. coli of Rhizopus oligosporus CJCC02-20 strain using a paper disk assay.

Hereinafter, the configuration and effects of the present application will be described in more detail through examples, but these examples are only exemplary descriptions of the present application, and the scope of the present application is not limited only to these examples.

Example 1. Isolation and identification of strains producing antibacterial substances

For the selection of strains with antibacterial activity, conventional tempes were cultured in PDA (Potato Dextrose Agar) medium for 5 days at 30° C. It was isolated by culturing for 5 days at 30°C in PDA medium containing ppm.

The nucleotide sequences of 18S rRNA, ABP (ABP1) and Chitinase (Chi2) genes of the strains selected by the above process were analyzed. For the nucleotide sequence, the GenBank database and each nucleotide sequence of the selected strains (18S rRNA: SEQ ID NO: 1, ABP1: SEQ ID NO: 2, Chi2: SEQ ID NO: 3) were compared using BLAST of NCBI.

Specifically, primers SEQ ID NOs: 4 and 5 were synthesized based on the nucleotide sequence of the ABP1 gene, and primers SEQ ID NOs: 6 and 7 were synthesized based on the nucleotide sequence of the Chi2 gene to confirm each nucleotide sequence through PCR. As a result, the nucleotide sequence of this strain showed 100% homology to Rhizopus oligosporus 18S rRNA, 99.9% to ABP Gene, and 100% homology to Chitinase Gene.

The strain obtained in this application from the above results was identified as a Rhizopus oligosporus strain and named as Rhizopus oligosporus CJCC02-20, and was transferred to the Korea Center for Microorganisms Conservation (KCCM), an international depository under the Budapest Treaty, in December 2020. It was deposited on the 09th and was given an accession number KCCM12893P.

Example 2. Isolation and Purification of Antimicrobial Substances

In order to isolate the antibacterial material produced by the newly isolated Rhizopus oligosporus CJCC02-20 strain, 10-15 g of the strain was inoculated into wheat bran, and then cultured at 30°C for 5 days and at 35°C for 1 day. The culture was diluted 50 times by weight of distilled water and then extracted at 30° C. and 150 rpm for 60 minutes. The crude extracts were filtered under reduced pressure with 5.0 μm filter paper using diatomaceous earth. The filtrate was freeze-dried at -20°C for 48 hours and purified to dryness.

Example 3. Antibacterial activity test against fungi and food poisoning-causing bacteria

In order to check whether the newly isolated strain of Rhizopus oligosporus CJCC02-20 has an inhibitory effect on fungi and food poisoning-causing bacteria, the following antibacterial activity test was conducted.

Aspergillus oryzae, a paste fermenting mold, and Aspergillus niger, a food contaminant, were cultured in PDA (Potato dextrose agar) medium.

The pathogenic yeast Candida albicans and the food contaminant Zygosaccharomyces bailii were cultured in YPD medium (Yeast extract peptone dextrose agar).

In addition, food poisoning-causing bacteria Bacillus cereus, enteropathogenic Escherichia coli, enteric fermentation bacteria Bacillus subtilis, Bacillus velezensis (Bacillus velezensis) are TSA (Tryptic soy agar) Cultured in medium.

After dissolving the dried antibacterial material in distilled water in Example 2, a paper disk of 8 mm diameter is placed on the plate medium on which the indicator strain is smeared, and 30 μl of the antibacterial material is constantly added thereto at 30° C. for 24 to 48 hours. By culturing, it was observed whether or not growth-preventing rings were generated by the antibacterial material produced by the isolated strain, and the results are shown in Table 1 and FIG. 1 . '-' in Table 1 is a case in which growth is not inhibited, in the case of '+', the diameter of the growth-stopping ring is 3 mm to 5 mm or less, and in the case of '++', the diameter of the growth-stopping ring is 6 mm to 10 mm. And, in the case of '+++', the diameter of the growth-stopping ring is 10 mm or more.

In addition, in addition to the above Rhizopus oligosporus CJCC02-20 strain, 4 types of Rhizopus oligosporus strains, which are strains isolated from conventional tempeh (tempe prepared in a traditional way in Indonesia and Vietnam), were tested for antibacterial activity in Comparative Examples 1 to 4. compared.

microbe antibacterial activity Rhizopus oligosporus CJCC02-20 Rhizopus oligosporus (Comparative Example 1) Rhizopus oligosporus (Comparative Example 2) Rhizopus oligosporus (Comparative Example 3) Rhizopus oligosporus (Comparative Example 4) Aspergillus oryzae +++ - - - - Aspergillus niger +++ - - - - Escherichia coli +++ - - - - Bacillus cereus ++ + + + + Bacillus subtilis + + + + + Bacillus velezensis + + + + + Candida albicans - - - - - Zygosaccharmyces bailii - - - - -

As can be seen in Table 1 and Figure 1, the purified material obtained from the Rhizopus oligosporus CJCC02-20 strain did not have antibacterial activity in yeast, but Aspergillus duck material and Aspergillus, which are difficult to inhibit in soybeans or general foods. It showed excellent antifungal activity to selectively control snigger mold bacteria.

In addition, it showed selective antibacterial activity against enteropathogenic E. coli, and it was confirmed that the antibacterial activity against Bacillus cereus, a food poisoning bacterium, was excellent compared to other Rhizopus oligosporus strains.

Example 4: Confirmation of the effect of controlling food poisoning bacteria in the fermented product tempe

In order to confirm the effect of controlling food poisoning bacteria in the fermented product of Rhizopus oligosporus CJCC02-20 identified in Example 1, 15-20 g of Rhizopus oligosporus CJCC02-20 strain was inoculated into wheat rice, and then at 30°C. Incubated for 6 days at 40°C for 1 day. The culture was made into a powder using a grinder.

In order to develop fermented products with diversified grains according to the consumption trend that changes every year, the above-mentioned culture solution powder and the commercially available Rhizopus oligosporus strain are added to steamed soybean, steamed soybean and rice, steamed soybean and wheat flour, and steamed wheat flour. (Ragi Tempe Raprima, Bandung Corporation, Indonesia) was inoculated and mixed with 4%, and then the humidity was adjusted to 60% and cultured at 30° C. for 1-2 days to make tempe, a fermentation product.

Cultured tempeh and commercially available conventional tempeh were cultured in Bacillus cereus selective medium supplemented with TSA (Tryptic soy agar), Polymyxin B and Trimethoprim for 1 day at 30°C. In the tempes inoculated with the Rhizopus oligosporus CJCC02-20 strain isolated in Example 1, Bacillus cereus and Escherichia coli, a food poisoning-inducing bacterium, were not detected, whereas the commercially available strains were not detected. It was confirmed that Bacillus cereus was detected in the inoculated tempeh and the conventional tempeh (see Table 2).

item conventional tempeh Tempe utilizing commercially available strains Tempe utilizing isolate strains Big head Soybean + Wheat Flour Soybean + Rice wheat flour Big head Soybean + Wheat Flour Soybean + Rice wheat flour Protease activity
(unit/g) 300 82 103 83 118 114 134 113 145 Bacillus cereus (CFU/g) 100,000 30,000 10,500 10,200 40,000 ND ND ND ND Escherichia coli (CFU/g) 10 ND ND ND ND ND ND ND ND

* ND: Not Detected

As a result of the above, it was confirmed that the Rhizopus oligosporus CJCC02-20 strain is a tempeh fermented strain and has an excellent effect in controlling food poisoning bacteria while implementing traditional flavor.

Therefore, the antibacterial composition comprising the novel Rhizopus oligosporus CJCC02-20 strain of the present application, the spores of the strain, the culture of the strain, its concentrate and its dried product, etc. has excellent antibacterial effect against fungi and food poisoning bacteria. It can be effectively used for food, quasi-drugs, and pharmaceuticals containing them.

From the above description, those skilled in the art to which the present application pertains will understand that the present application may be embodied in other specific forms without changing the technical spirit or essential characteristics thereof. In this regard, it should be understood that the embodiments described above are illustrative in all respects and not restrictive. The scope of the present application should be construed as including all changes or modifications derived from the meaning and scope of the claims to be described later rather than the above detailed description and their equivalent concepts to be included in the scope of the present application.

Korea Microorganism Conservation Center (Overseas) KCCM12893P 20201209

<110> CJ CheilJedang Corporation <120> Novel Rhizopus oligosporus strain and antibacterial and antifungal composition comprising the same <130> KPA201759-KR <160> 7 <170> KoPatentIn 3.0 <210> 1 <211> 1746 <212> RNA <213> Unknown <220> <223> Rhizopus oligosporus CJCC02-20 18s rRNA <400> 1 caagattaag ccatgcatgt ctaagtataa ataactttat attgtgaaac tgcgaatggc 60 tcattaaatc agttatgatc tacgtgacaa attctttact acttggataa ccgtggtaat 120 tctagagcta atacatgcaa aaaagccctg actcacgaag gggtgcactt attagataaa 180 accaacgcgg ggtaaaacct gtttcgtggt gaatcataat aattaagcgg atcgctggcc 240 ttgagccggc gacggtccac tcgattttct gccctatcat ggttgagatt gtaagataga 300 ggcttacaat gcctataacg ggtaacgggg aattagggtt cgattccgga gagggagcct 360 gagaaacggc taccacatcc aaggaaggca gcaggcgcgc aaattaccca atcccgacac 420 ggggaggtag tgacaataca taacaatgca gggcctttac ggtcttgcaa ttggaatgag 480 tacaatttaa tcccttaacg aggatcaatt ggagggcaag tctggtgcca gcagccgcgg 540 taattccagc tccaatagcg tatattaaag ttgttgcagt taaaacgtcc gtagtcaaac 600 tttagtctta ccggcgcggt ggcctggtct tcattgatca agctcgctgt cgctggagac 660 tccacgtccg ctggctccta gtcctcgtgg ctagggtttt gtggacaatt accatgagca 720 aatcagagtg tttaaagcag gctttaagct tgaatgtgtt agcatggaat aatgaaatat 780 gactttagtc ctattttcgt tggtttaggt actatagtaa tgattaatag aaacggttgg 840 gggcatttgt atttgcccgc tagaggtgaa attcttggat tggccgaaga caaactactg 900 cgaaagcatt tgacccagga cgttttcatt gatcaaggtc taaagttaag ggatcgaaga 960 cgattagata ccgtcgtagt cttaaccaca aactatgccg actagagatt gggcgtgttt 1020 cttttgactc gctcagcatc ttagcgaaag taaagttttt gggttctggg gggagtatgg 1080 gacgcaaggc tgaaacttaa aggaattgac ggaagggcac caccaggagt ggagcctgcg 1140 gcttaatttg actcaacacg gggaaactca ccaggtccag acatagtaag gattgacaga 1200 ttgaaagctc tttctagatt ctatgggtgg tggtgcatgg ccgttcttag ttcgtgagtg 1260 atttgtctgg ttaattccga taacgaacga gaccttattc tgctaattac acaggctaac 1320 tctttcgggt tggtttatgt ttatcattta actgtgttct tagagagact atctgcttga 1380 agcagaggga agttttaggc aataacaggt ctgtgatgcc cttagatgtt ctgggccgca 1440 cgcgcgctac actgatgaag tcagcgagtt ataaccttgg ccggaaggtc tgggtaaact 1500 tttgaaactc atcgtgctgg ggatagagca ttgtaattat tgctcttcaa cgaggaattc 1560 ctagtaagcg cgagtcatca gctcgcgttg attacgtccc tgccctttgt acacaccgcc 1620 cgtcgctact accgattgaa tggttatagt gagcatatgg gatccgcaga atttgactgg 1680 caacagtcat tttctgcaga gaactatggc aaactaggct atttagagga agtaaaagtc 1740 gtaaca 1746 <210> 2 <211> 1268 <212> DNA <213> Unknown <220> <223> Rhizopus oligosporus CJCC02-20 ABP1 gene <400> 2 agctgtagtg caatgttcaa gctaaaaata agtttgtgta tgtgctctgc aactgcatga 60 tattctagaa cgactgtgtt gcattttctt ttaattgatc tgtttgttgg tttctagaag 120 taatgcaaaa acggttgtac aaacatacaa aaggtcatat gacagatacg ccatcctaat 180 aagcgaaata gtgagcacct gcaagggcat tttaattaat aaagtgactt aatagcaatt 240 atggggcttt caagcttcca gggggtattt tgtttattgc ttaagaaaag tgctgtcctt 300 gtattcaaaa gtctatattt ttttgttata taaatataca catggatcac taaactattt 360 cttggaggta cagagaaaga gaggggaagg aagaacacga tccagcatca tggtcgctat 420 ggtgtcactt cgtcaacaga cgtcatttta gctcaccctt ctctcctttc tctttttctc 480 gcttgtcttc cttttttagt aatgttatac atttaagcgg tttatgtggt accttgccat 540 ttacataaat aaggaagctc aagattttgc aagatgctag ttctttctca aatgaagttg 600 atctacgttt tttgtttctt tactttgctc ttaattgcta ctcaacaagt gagtgctgca 660 tgtggtgctg aaggttcatg tcatggtttc ggaggcggag agttgtgtaa tgacagatgc 720 aaacgatgct ctggccctac tggaaaatac aaacggggag cctgttgtgg tactttgaaa 780 caagcttgct gctgctatta cagctaatta atgatgtgat gtgtttaact ttataaagca 840 gttgctttat aaaattgaaa attgaaaaat aaaaatgtct gtctccactc ctctgtcttc 900 ttgtttgggg ttttaattca tatgtagacg tgttacatgc tctaattatc aattacagca 960 aatgatcatc aagaggcagc agggtgtaat gaaaaacaca agtccaacac ttagagagct 1020 cattttttgc gtttttgcgc ctcaaatgaa atgcaataat ttcgttcaat caaagctttt 1080 tttggatttt aatatgtaca agattgtaca gccattatac tattacaatc aaataaagcc 1140 tatttctctt ttcttgtttc ttctatttaa gcatttccaa cattcaaaag caagttagag 1200 ccttcgccct taccactagg taaataagcc actttggagg tttgtgataa tgtctgtgca 1260 atatcctt 1268 <210> 3 <211> 2119 <212> DNA <213> Unknown <220> <223> Rhizopus oligosporus CJCC02-20 chi2 gene <400> 3 gcatgcgcat agtaaacaat gcttaaacca gatgacatac atacgtctat gaatgcaact 60 gattgatcat gaaatagccg cttctttgtt ttcgaatgat atccctacca tttcttcctt 120 tttatttcaa tgtaacatga gttttaatag gtaatatatt acttgatatt attacttgat 180 attacctatg accctaatct tattcattaa cgtgtaactt gagctgcaat gggaaaaaaa 240 aagataatga agacatggca acattagccc acgcatataa atactcactg gacatcctga 300 catcaaatca ttatctttac acttttcttt cccttattct tcaacatgct cactcgtact 360 ttccttggaa tggctatatc tgccttcttg gcatcaactg gtgttcaagc tgcttggtcc 420 tctcatggcc ctaatgtcat gtactactgg ggtcaaaatt ctgctggcgg atcaaataca 480 caagcatcct taggcactta ctgtgagtct ggccaagtcg atgccgtcct cctgtccttc 540 cttcacgtct ttaacgttgg tggtatccct gaaatcaatc tttctagcgc ctgtgctggt 600 acctattttc ccaatactca actacttagt tgtcctgctg ttggtgctgg taagtgttaa 660 accttttgtt gtcggggtct cttaacatct ctttcttgat agatatcaag aaatgtcaag 720 acaagggcgt caaggtcata ctctctcttg gtggtgctgc tggtgtctat ggtttcacaa 780 gcgatgccca aggacaacag ttcgcacaga cgatctggaa cctctttggt ggtggaaact 840 cagacacacg tccatttggt gatgctgtta ttgacggtgt cgatcttgac attgaaggtg 900 gctcatctac gggttatgtc gcctttgtga atgctcttcg tcaaaagttc tccagcaact 960 tccttattgg tgctgctcct caatgcccct tccctgacgc catcctcggc agtgtgttga 1020 actcggctag tttcgactac gtcaatgttc agttctataa caactattgc tctgctactg 1080 gttcctcctt caactttgac acctgggata actgggccaa gacgacgtca cctaacaaga 1140 acgtcaagat catgtttacg gttcctggct catctactgc tgcaggcagc ggctacgttc 1200 ccatgtctac tcttcaaact atagttcctt ctcttgcttc caagtactcc agctacggcg 1260 gtgtctcagt ctgggatgcc tctcaagcct ggaataacgg tggcttcaac tctcaactct 1320 actcgcttgt ccacagcggt ggctccactc ctcctcctcc ttcctcttct tcagctacca 1380 agacaacaac aaaaacgaca gcaacaagca ccaagacgac gactacgact gcccctactg 1440 ctaccagcac acctggcagt tgtcctgttg ccaatcagcc ttgctctact caaaatcaat 1500 atgcttgtac agccgatggc aagtacgctg tctgcgacca tggcaaatgg gttgcctcct 1560 cctgcccctc caacactgtc tgcattccta ctaccgatgg tgcctctatc tactgtggtt 1620 atgctactgg ctctggcagc acttgtcctt cagtcagcgc actcgaaatt acagctgctt 1680 ctctcggctc taagaatggc cccgtacctc gcccatacaa ggcttccaag gttgctgctc 1740 aactcgctgt tacttccact gacaagaata gctttgaagc tgttatcaat gctcgtcgca 1800 ccacgctcac tccctttgaa aaatctgtta ctatcgaatt cactacacct tccaacatca 1860 agtttaccga atctgatatg ggtcccgttc gtcaagtcgg caacaaggtt cgtattcaag 1920 ccaagaatga ttataacgaa tccatgactc tcgtcgtcaa agtcaagggt tccatcaatt 1980 caggcgtctt tgtggctccc agtacttctg cttggaactt taaataaata ttgtattaga 2040 taataataat aataaaaaaa gtgcaataac gttttattgt acgtattgaa gcagatttat 2100 gtgtattata ttattaaaa 2119 <210> 4 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> ABP1 gene reading pair forward primer <400> 4 gagaaagaac tagcatcttg c 21 <210> 5 <211> 16 <212> DNA <213> Artificial Sequence <220> <223> ABP1 gene reading pair reverse primer <400> 5 catcattaat tagctg 16 <210> 6 <211> 23 <212> DNA <213> Artificial Sequence <220> <223> chi2 gene reading pair forward primer <400> 6 caacatgctc actcgtactt tcc 23 <210> 7 <211> 21 <212> DNA <213> Artificial Sequence <220> <223> chi2 gene reading pair reverse primer <400> 7 gagccacaaa gacgcctgaa t 21

Claims (14)

Rhizopus oligosporus CJCC02-20 (Rhizopus oligosporus CJCC02-20) strain deposited with accession number KCCM12893P with antibacterial activity against fungi and bacteria,
The fungus is Aspergillus ( Aspergillus ) genus, the bacteria is Bacillus cereus ( Bacillus cereus ) or Escherichia coli ( Escherichia coli ) Will, the strain.
delete The strain of claim 1, wherein the fungus is Aspergillus oryzae , Aspergillus niger , or a combination thereof.
delete The strain according to claim 1, wherein the strain has a specific antibacterial activity against Aspergillus duck material, Aspergillus niger or E. coli.
[Claim 6] The Rhizopus oligosporus CJCC02-20 strain according to any one of claims 1, 3 and 5, the spores of the strain, the culture of the strain, a concentrate thereof, and a dried product thereof selected from the group consisting of An antibacterial composition comprising any one or more as an active ingredient.
delete A food composition comprising the antimicrobial composition according to claim 6 .
The food composition according to claim 8, wherein the food is tempeh or a sauce comprising tempeh.
A feed composition comprising the antimicrobial composition according to claim 6 .
A pharmaceutical composition for the prevention or treatment of microbial infections caused by fungi, bacteria, or a combination thereof, comprising the antimicrobial composition according to claim 6 .
The pharmaceutical composition of claim 11, wherein the microbial infectious disease is any one or more selected from the group consisting of food poisoning, diarrhea, vomiting, enteritis, gastroenteritis, constipation, abdominal pain and abdominal distension.
A health functional food composition for preventing or improving microbial infections caused by fungi, bacteria, or a combination thereof, comprising the antibacterial composition according to claim 6 .
A quasi-drug composition comprising the antimicrobial composition according to claim 6 .

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